Proinsulin maturation, misfolding, proteotoxicity

Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical Center, Ann Arbor, MI 48109, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 11/2007; 104(40):15841-6. DOI: 10.1073/pnas.0702697104
Source: PubMed


As a tool to explore proinsulin (PI) trafficking, a human PI cDNA has been constructed with GFP fused within the C peptide. In regulated secretory cells containing appropriate prohormone convertases, the hProCpepGFP construct undergoes endoproteolytic processing to CpepGFP and native human insulin, which are specifically detected and cosecreted in parallel with endogenous insulin. Expression of C(A7)Y mutant PI results in autosomal dominant diabetes in Akita mice. We directly identify the misfolded PI in Akita islets and also show that C(A7)Y mutant PI, either in the context of the hProCpepGFP chimera or not, engages directly in protein complexes with nonmutant PI, impairing the trafficking and recovery of nonmutant PI. This trapping mechanism decreases insulin production in beta cells. Thereafter we observe a loss of beta cell viability. The data imply that PI misfolding leading to impaired endoplasmic reticulum exit of nonmutant PI may be a key early step in a chain reaction of beta cell dysfunction and demise leading to onset and progression of diabetes.

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    • "Insulin biosynthesis is strongly dependent on posttranscriptional regulatory mechanisms, including the modulation of translation and stability [5] [6] [7]. Most of the mutations with INS gene are predicted to disrupt the folding of the proinsulin molecule. "
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    ABSTRACT: Neonatal diabetes mellitus (NDM) is a rare condition with a prevalence of 1 in 300,000 live births. We have found 3 known SNPs in 5'UTR and a novel SNP in 3' UTR in the INS gene. These SNPs were present in 9-month-old girl from Saudi Arabia and also present in the father and mother. The novel SNP we found is not present in 1000 Genome project or other databases. Further, the newly identified 3' UTR mutation in the INS gene may abolish the polyadenylation signal and result in severe RNA instability. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
    Diabetes research and clinical practice 07/2015; 109(3). DOI:10.1016/j.diabres.2015.06.002 · 2.54 Impact Factor
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    • "Betacells are professional secretory cells, which make extensive use of the ER machinery to synthesize insulin and other proteins (Alberts et al., 2002; Ghaemmaghami et al., 2003; Rutkowski and Kaufman, 2004; Schnell, 2009). Insulin is synthesized in the ER at a rate of 1 million molecules per minute (Anelli and Sitia, 2008; Liu et al., 2007; Schuit et al., 1988). It has been suggested that mild persistent hyperglycemia and inflammatory signaling, e.g., through IL-1, can trigger ER-stress (as well as oxidative stress and mitochondrial stress, respectively) and beta-cell apoptosis (Atkinson et al., 2011; Marchetti et al., 2007). "
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    ABSTRACT: In type 1 diabetes, an autoimmune disease mediated by autoreactive T-cells that attack insulin-secreting pancreatic beta-cells, it has been suggested that disease progression may additionally require protective mechanisms in the target tissue to impede such auto-destructive mechanisms. We hypothesize that the autoimmune attack against beta-cells causes endoplasmic reticulum stress by forcing the remaining beta-cells to synthesize and secrete defective insulin. To rescue beta-cell from the endoplasmic reticulum stress, beta-cells activate the unfolded protein response to restore protein homeostasis and normal insulin synthesis. Here we investigate the compensatory role of unfolded protein response by developing a multi-state model of type 1 diabetes that takes into account beta-cell destruction caused by pathogenic autoreactive T-cells and apoptosis triggered by endoplasmic reticulum stress. We discuss the mechanism of unfolded protein response activation and how it counters beta-cell extinction caused by an autoimmune attack and/or irreversible damage by endoplasmic reticulum stress. Our results reveal important insights about the balance between beta-cell destruction by autoimmune attack (beta-cell homicide) and beta-cell apoptosis by endoplasmic reticulum stress (beta-cell suicide). It also provides an explanation as to why UPR may not be a successful therapeutic target to treat type 1 diabetes.
    Journal of Theoretical Biology 05/2014; 375. DOI:10.1016/j.jtbi.2014.05.003 · 2.12 Impact Factor
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    • "This prediction is supported by diabetic mouse models of misfolded proinsulin, namely the Akita (Ins2WT/C96Y) mouse15 and Munich (Ins2WT/C95S) mouse3. In both models, the mutant insulin interferes with secretion of wild‐type insulin, and β‐cell mass is decreased. "
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    ABSTRACT: Aims/Introduction The human insulin gene/preproinsulin protein mutation C43G disrupts disulfide bond formation and causes diabetes in humans. Previous in vitro studies showed that these mutant proteins are retained in the endoplasmic reticulum (ER), are not secreted and are associated with decreased secretion of wild‐type insulin. The current study extends this work to an in vivo zebrafish model. We hypothesized that C43G‐green fluorescent protein (GFP) would be retained in the ER, disrupt β‐cell function and lead to impaired glucose homeostasis. Materials and Methods Islets from adult transgenic zebrafish expressing GFP‐tagged human proinsulin mutant C43G (C43G‐GFP) or wild‐type human proinsulin (Cpep‐GFP) were analyzed histologically across a range of ages. Blood glucose concentration was determined under fasting conditions and in response to glucose injection. Insulin secretion was assessed by measuring circulating GFP and endogenous C‐peptide levels after glucose injection. Results The majority of β‐cells expressing C43G proinsulin showed excessive accumulation of C43G‐GFP in the ER. Western blotting showed that C43G‐GFP was present only as proinsulin, indicating defective processing. GFP was poorly secreted in C43G mutants compared with controls. Despite these defects, blood glucose homeostasis was normal. Mutant fish maintained β‐cell mass well into maturity and secreted endogenous C‐peptide. Conclusions In this model, the C43G proinsulin mutation does not impair glucose homeostasis or cause significant loss of β‐cell mass. This model might be useful for identifying potential therapeutic targets for proper trafficking of intracellular insulin or for maintenance of β‐cell mass in early‐stage diabetic patients.
    03/2013; 4(2):157-67. DOI:10.1111/jdi.12015
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